Regulation of actin dynamics plays an integral role in many processes important for human health. Morphogenesis during embryonic development involves extensive migration, establishment of cell: cell contacts, regulated secretion and internalization of signaling proteins and numerous other cellular processes dependent upon dynamic actin remodeling. Disease-related processes including metastatic cancer, inflammatory disorders, wound repair and tissue regeneration all involve actin-dependent mechanisms. Cell migration requires coordinated regulation of cellular protrusions, adhesion, contractile forces and rear detachment. Ena/VASP proteins regulate the protrusive step of motility in fibroblasts by controlling the geometry of actin networks within lamellipodia, promoting formation of longer, less branched actin networks. Ena/VASP also likely play key roles in other actin-dependent processes such as phagocytosis, formation of cell: cell junctions and regulation of the vascular endothelium. We hypothesis that Ena/VASP proteins act as key convergence points, integrating specific signals and converting them into dynamic cytoskeletal remodeling. This grant outlines experiments to identify other migration and actin-dependent processes that utilize Ena/VASP function in vivo through analysis of mice lacking two or all three family members. W will also combine cell biological and biochemical approaches to continue our efforts to elucidate mechanisms of Ena/VASP function and regulation. We will also characterize interactions between Ena/VASP and NESH, a member of the Abi family of adaptor molecules that localizes to protruding lamellipodia and filopodia. Interestingly, Abi proteins are present in complexes that regulate the SCAR/WAVE family of Arp2/3 activating proteins, suggesting that NESH could serve as a link between Ena/VASP and SCAR/WAVE function.